The control of multidrug resistant tuberculosis has been severely hampered by the lack of rapid methods for antimicrobial susceptibility testing and diagnosis of mycobacterial infection. The research proposed provides a novel approach to rapid antibiotic susceptibility testing, based on One Cell Systems' proprietary microdrop technology. Individual mycobacterial microcolonies will be encapsulated and grown within a very small cloning region defined by a gel microdrop (GMD). Automated analysis of GMDs by flow cytometry following clonal growth and fluorescent staining of the organisms forms the basis of this assay system. Susceptibility of mycobacteria to antibiotics has been detected in 5 days using the GMD- clonal growth method, thus demonstrating the basic feasibility of this approach. The goals of the Phase I research are to develop quantitative methods for evaluation of mycobacterial growth and resistance to antimicrobial agents. These experiments will form the basis for a commercially viable, rapid mycobacterial assay system, utilizing conventional methods at a microscale. The re-emergence of tuberculosis as a public health problem in the United States points to the urgent need for such a testing system.